Skin is a large organ, crucial for life, which protects the organism against environmental stresses, such as physical, chemical and mechanical stresses, and which prevents water loss. The complexity of skin notably arises from the association various tissues having different embryologic origins. While the barrier functions of skin depend on the epidermis, through the differentiation process of keratinocytes, skin homeostasis depends on the balance of multiple cellular and tissular interactions in which the dermis plays a key role.
Immediately after a skin injury has occurred, several events take place to repair the damaged tissue. Wound healing is a complex and dynamic process involving soluble mediators, blood cells, extracellular matrix components, and resident cells, including fibroblasts.
Briefly, the wound healing process includes three interactive phases: inflammation, granulation tissue formation and remodelling. This sequence of events aims at the recovering of tissue integrity and the restoration of its functions. The quality of the healing—which should ideally lead to an absence of scar and to the reestablishment of tissular function—is thus depending on a complex equilibrium.
In many species, the fetus possesses the unique ability to heal skin wounds without scar formation (Estes et al, (1994) Differentiation 56:173; Ferguson et al. (1996) Plast. Reconstr. Surg. 97:854), nevertheless adults will always present aftereffects that can lead to functional disorders.
In some cases, problems occur during the wound healing process, leading to excessive scar development consecutive to an excessive extracellular matrix deposition (e.g. hypertrophic scars and keloids). In contrast, a disturbance in wound healing may also be characterized by poor healing or an absence of healing, as is notably observed in diabetes, and pressure, arterial or venous ulcers.
Hypertrophic scars and contraction features following burn wounds lead to functional troubles. The treatment for such wounds generally consists in the use of compressive dressings that will have to be kept during months or years. In most cases additional surgical operations will be necessary.
Furthermore, excessive scarring, poor healing or absence of healing, affect the quality of life of patients, but also represent a high cost.
Various treatments are currently used to try to restart the wound healing process in chronic wounds, such as the use of detersion, of growth factors, or of vacuum therapy. However, in many cases these treatments have proven unsatisfactory.
Thus, improvement of the functionality of “healed” area after wound closure is a first objective in the management of skin wounds. Besides, promotion and stimulation of the wound healing process in chronic wounds is another objective.
As such, grafting of cells or grafting of in vitro reconstructed tissues appears to be a promising field in the treatment of skin wounds.
In the frame of skin wound healing, the minimum requirement is to re-establish a barrier function to avoid infection and water loss. It is the horny layer of the epidermis (the product of terminal keratinocyte differentiation) that plays this role. However, although the barrier function depends on the epidermis, there is also a need to improve grafting by incorporating dermal tissue in order to promote the functionality of the engrafted zone.
Dermis neo-formation is an important step in wound healing since dermis accounts for a number of the mechanical properties of skin and promotes the formation and anchoring of a neo-epidermis, in particular through the activation of growth and differentiation of keratinocytes.
Dermal fibroblast grafting in the frame of the management of skin wounds has been shown to accelerate the formation of a neo-dermis and to improve the functionality of the grafted area (Coulomb et al. (1998) Plast. Reconstr. Surg, 101:1891-1903). This is notably due to the promotion of the synthesis of elastin which contributes to the mechanical properties of the skin. This improvement in dermis directly impacts on the organization and anchorage of epidermis.
Thus, in humans, a normal undulated dermo-epidermal junction is formed within one year of dermal fibroblast grafting in a skin wound whereas, in the absence of such a grafting, from 3 (in children) to 5 years (in adults) are required to obtained the same result.
However, the quality of wound healing, i.e. disappearance of wound marks (scar) and reestablishment of the functional properties of skin, is the result of a delicate balance and most of the time after-effects can not be prevented.
In addition, in situations such as large burns, dermal fibroblasts cannot be available in sufficient number.
As such, it is an object of the present invention to provide an advantageous alternative to the use of dermal fibroblasts in the management of skin wounds.
Other fibroblasts have been explored for their implication in wound healing. However, they have been found less efficient than dermal fibroblasts. For instance, fibroblasts of the adipose tissue although being similar to dermal fibroblasts in many respects (van der Bogaerdt et al. (2002) Arch. Dermatol. Res. 294:135-142) have proven less efficient than dermal fibroblast in promoting growth and differentiation of keratinocytes for the formation of an epidermis (Middelkoop (2005) Int. J. Low Extrem. Wounds 4:9-11).
Gingival fibroblasts are mesenchymal cells which are capable of migrating, adhering and proliferating within the soft connective tissues of the gum, thereby maintaining the integrity of the gingival tissue which is exposed to numerous aggressions, such as mechanical stresses, bacterial infections, or pH and temperature variations. Gingival fibroblasts are in particular described in Gogly et al., (1997) Clin. Oral Invest. 1:147-152; Gogly et al. (1998) Biochem. Pharmacol. 56:1447-1454; and Ejeil et al. (2003) J. Periodontal. 74:188-195.
Depending on environmental conditions, gingival fibroblasts are capable to modulate their phenotype, and to respond by proliferating, migrating, synthesising matrix components or matrix-related enzymes.
Gingival fibroblasts synthesise collagens (e.g. types I, III, V, VI, VII, XII) elastic fibers (oxytalan, elaunin and elastin), proteoglycans and glycosaminoglycans (e.g. decorin, biglycan), glycoproteins (e.g. fibronectin, tenascin). Simultaneously, gingival fibroblasts synthesise enzymes that are able to degrade the macromolecular compounds (matrix metelloproteinases; MMPs), but also enzymes inhibiting active forms of MMPs (Inhibitors of metalloproteinases; TIMPs). Gingival fibroblasts are thus important actors of extracellular matrix remodelling.